Simple, sensitive, and label-free miRNA analysis through strand displacement reaction integrating with G-quadruplex-based signal generation

Abstract MicroRNAs (miRNAs) play a significant role in the pathogenesis of various diseases throughout biological processes, and the accurate detection of miRNA biomarkers holds great potential for early stage disease diagnosis and treatment. In this study, a novel method is developed to detect miRNA-21, a biomarker for drug-induced liver injury, by combining target sequence recycling with G-quadruplex-based signal production. This approach is highly sensitive and does not require the use of labels. The target sequence facilitates the cyclic exposure of G-rich regions in the detection probe by toehold-mediated strand displacement processes, with the aid of the catalytic chain. The G-quadruplex sequences that have been produced subsequently interact with thioflavin T (ThT), resulting in a significant increase in its fluorescence intensity. This enhanced fluorescence is utilized for the purpose of detecting miRNA-21, with a remarkably low detection limit of 4.4 fM. The suggested technique also allows for the very specific identification of the target miRNA-21. Due to its non-label format, excellent selectivity, and sensitivity, this technology presents a straightforward and versatile approach for detecting a wide range of biomarkers in the early phases of illness detection.